ABSTRACT The cervical spinal cord (CSC) is extremely compact: all motor and sensory tracts pass through a small cross-sectional area. Thus, damage in the CSC often leads to severe disability and can be devastating. Yet clinicians lack sensitive tools for diagnosing and grading CSC damage, so treatment and outcomes vary widely. A more sensitive tool would greatly improve the characterization of CSC injury in patients with multiple sclerosis (MS) and cervical spondylotic myelopathy (CSM). In MS patients, for instance, there is no way to know if symptoms are due to reversible demyelination (potentially treatable with drugs) or untreatable, permanent axonal damage. In CSM patients, surgery is a viable option if axons are preserved, but there are no validated techniques to determine this. Although the diffusion tensor MRI (DTI) has emerged as a promising method to quantitatively evaluate CSC pathology, DTI lacks the specificity about pathological information. For instance, increased radial diffusivity may be induced by either/both increased extra-axonal water contents and demyelination. Therefore, we have been developing a new imaging method called ultra-high B Diffusion-Weighted MRI (UHb- DWI) that promises to distinguish demyelination, inflammation, and axonal damage. This method selectively suppresses the signal contribution from extra-axonal compartment in order to isolate the signal of intra-axonal water. Our method includes: 1) a novel data acquisition, 2) custom MRI RF coils, and 3) Monte-Carlo simulation (MCS) of water diffusion in white-matter fiber. Using these tools, we are observing very strong evidence that UHb-DWI provides information, specific to the degree of myelination in the spinal cord. A confounding factor discovered in our preliminary data on healthy subjects is that the UHb-DWI signal behaves differently for motor and sensory tracts, and also on the age of subjects. Since the MCS predictive model cannot be applied uniformly, it is clear that our method requires a reference library of diffusion metrics from healthy CSCs. Also, to validate that UHb-DWI is actually detecting injury due to demyelination and axonal damage, we seek to correlate UHb-DWI metrics with gold standard ex-vivo histopathology of animal model of spinal cord injury. Therefore, the goals of this study are to: (a) validate that biomarkers measured with UHb- DWI can detect demyelination and axonal loss in an animal model of cord injury, (b) establish a reference library of healthy human CSC data in eight age/gender groups, and (c) acquire exploratory data on a limited cohort of MS patients to establish proof of concept for human applications. Upon successful completion of current work, we will have a powerful, non-invasive imaging method to characterize axonal density and demyelination in patients with spinal cord injury. This may lead to earlier and more sensitive detection of clinically important spinal cord lesions or the ability to monitor the evolution of the cord during the treatment of patients with MS, CSM and other spinal cord diseases.